Effect of different centrifugationduration on Simmental bull sperm quality and membrane status after sexing, cooling and freezing processes

The Percoll density-gradient centrifugation was the most widely used method that able to affect sperm quality during the separation of X and Y spermatozoa. Sexing using Percoll density-gradient centrifugation method at 850 x g for 5 min could afford 85% of X and Y sperm separation. However, the technical problem would appear for 5 min centrifugation in which sperms at the under layer population were not able to obtain the tube base and led to less X and Y sperm separation. Whereas the treatment for 7 min centrifugation had a high level separation power as sperms were able to reach the tube base. The study examines the effect of different duration of Percoll density-gradient centrifugation method on the quality of spermatozoa after sexing, before freezing, and post thawing processes. Fresh semen sample were routinely collectedtwice a weekfrom two Simmental bullsaged4-9years usingartificialvagina at Ungaran artificialinsemination (AI) center, Central Java, Indonesia. The sperm which hadindividualmotility more than 70% were selected forthe freezing process. Sexing sperm using Percoll density-gradient centrifugation method for 5 and 7 min was applied in this study. Sperm quality after sexing, before freezing, and post thawing processes was assessed under a light microscope at400 xmagnification and its concentration was evaluated using haemocytometer. Data were analyzed by a pair of t-test. For all statistical analyses, the level of significance was P<0.05. There were significant differences (P<0.05) on sperm qualities among 5 and 7 min of centrifugation treatments. The study demonstrated that 5 min centrifugation treatment using Percoll gradient provided significantly better sperm motility, viability and concentration compared to 7 min centrifugation treatment (P<0.05). Furthermore, a lower number of abnormal sperms were also found significantly at 5 min centrifugation treatment. Our findings suggest that 5 min would be a good and acceptable duration to centrifuge Simmental bull sperms using Percoll density-gradient.